Tile vast majority of cervical and anogenital cancers are caused by papillomavirus infection. This proposal focuses on optimization of a papillomavirus inhibitor and is submitted in response to PA-01-113 on AIDS-associated opportunistic infections and malignancies. HIV-infected individuals are particularly susceptible to developing papillomavirus infections and associated malignancies. Papillomavirus infection is also a major problem in the general population. This year 3 million women in tile U.S. will develop cervical dysplasia, the premalignant condition that can progress to cervical cancer. Although the viral genome and proteins are logical targets for the creation of inhibitors, no specific antiviral treatment yet exists. The key regulator of viral transcription and replication is the viral E2 protein, which has an N-terminal activation domain and a C-terminal DNA-binding/dimerization domain. When fused to a small peptide that mediates efficient cellular uptake, a repressor form of E2 that lacks the N-terminal activation domain is an inhibitor (ICs0 of 85 nM) of transcriptional activation in cell culture. The three-dimensional structure of the E2/DNA complex will be determined and used to guide refinements of the lead inhibitor along two tracks. One form of the E2 inhibitor is designed to bind and inactivate wild-type viral E2 protein through heterodimer formation; inhibition of viral E2 is expected to stop active viral infections. The second form of inhibitor is designed to target the E2 binding sites on viral DNA to repress viral E6 and E7 expression. E6 and E7 proteins allow viral propagation and predispose cells toward malignancy by stimulation of degradation and subsequent loss of p53 and Rb. Modified inhibitors will be evaluated for DNA affinity, dimerization, and stability, and will be assayed for their pharmacologic properties, including cellular uptake, half-life, and toxicity. The biochemical and pharmacologic properties of each inhibitor will be correlated with results from assays for inhibition of cell proliferation and viral replication. The optimal cellular delivery system for the best inhibitors will be defined. The overall goal is to achieve high potency in cell-based models for eventual testing in animals and in human clinical trials.